耐藥性細菌的迅速傳播極大的影響了臨床病原體的診斷與治療，且由於過度使用抗生素，細菌對抗菌劑的耐藥性已成為現今醫療保健的重要問題。其中，泌尿道感染是影響人類常見的細菌感染性疾病，尤其發生在女性的機率較高，且極大的影響腎臟功能。根據不同的感染部位，也會出現不同的症狀，此外，泌尿道感染具有很高的復發率。在臨床治療上，高劑量的廣效型抗生素常被用以抑制或殺死泌尿道感染的病原體，然而，殘留的存活細菌可能會產生耐藥性，這使得泌尿道感染在往後的治療更為棘手。為了減少廣效性抗生素的使用劑量，應在給藥前根據一種被稱為抗微生物敏感性試驗的常規方法確定抗生素的最小抑制濃度。然而，傳統的敏感性試驗在篩選抗生素劑量是相當耗時 (20小時 ~ 70小時) 且耗費人力的。為了縮短AST的周轉時間，將人為操作的步驟自動化是非常需要的。因此，本研究提出了一種自動化的微流體系統，它可以在人工干預最少的情況下執行AST程序。在這項研究中展示了一新型的便攜式檢測系統，該系統能夠依據序列肉湯稀釋法同步將兩種菌株進行兩種不同抗生素的敏感性試驗。透過此自動化平台，抗生素敏感性試驗流程的進行可以大量的減少人為干擾，並將檢測時間從20小時縮短為4個半小時至9小時。此外，透過比色測定法，可以經由圖像分析確定抗生素對細菌的最小抑制濃度。總結來說，此具前瞻性發展的檢測平台在未來可以實現基於抗微生物敏感性試驗之個人藥品的即時檢測。

The quick spread of antibiotic-resistant bacteria has greatly impacted clinical pathogen diagnosis and treatment. Due to the overuse of antibiotics, drug resistance has become a significant issue in modern health care. Among them, urinary tract infections (UTIs), which are common bacterial infections affecting humans, especially women, which greatly impact kidney functions. Symptoms vary based on different infection locations and bacterial loading. Furthermore, UTIs also have a high recurrence rate. In clinical treatment, UTIs are treated with high dosages of broad-spectrum antibiotics to inhibit or kill the pathogens. However, residual surviving bacteria may evolve drug resistance, which makes UTIs extremely tricky to be cured. To reduce the dosage of broad-spectrum antibiotics, the minimum inhibitory concentration (MIC) of antibiotics should be determined before administration based on a common practice referred to as antimicrobial susceptibility testing (AST). However, the conventional AST approach is relatively time-consuming (20hrs ~ 72hrs) and labor-intensive to screen antibiotic dosage. In order to shorten the turnaround time of AST, automating the manual process is of great need. This study therefore presents an automated microfluidic system which could perform AST with minimum human intervention. In this study, a new, portable system capable of conducting two different AST measurements on two different strains of bacteria with serial broth dilutions in parallel was demonstrated. Using this automated platform, the whole procedure of AST could be carried out with greatly reduced human interruption and therefore shorten the detection time from twenty hours to 4.5 ~ 9 hrs. Moreover, based on a colorimetric assay, the MIC values could be determined via image analysis. In summary, this device may be promising to realize point-of-care personalized medicine for AST in the near future.

Abstract............................1
中文摘要.............................3
Acknowledgements....................4
Table of Contents...................6
List of Figures.....................9
List of Table.......................15
Nomenclature and Abbreviations......16
Chapter 1 Introduction..............19
1-1 Bio-MEMS and microfluidic system...................19
1-2 Urinary tract infection ...........................21
1-3 Antimicrobial susceptibility test (AST)............23
1-4 Integrate microfluidic systems for antimicrobial susceptibility tests..................................................24
1-5 Point-of-care (POC) diagnostics....................25
1-6 Motivation and novelty.............................26
Chapter 2 Materials and Methods........................29
2-1 Entire experimental procedure......................29
2-2 The procedure of bacteria capturing by using nano-magnetic beads..................................................32
2-3 Bacteria quantification............................34
2-4 Pneumatically driven microfluidic devices..........36
2-5 Portable modules for automatically performing the on-chip AST .......................................................41
2-5-1 Pneumatic control module and electromagnetic valve control module.................................................41
2-5-2 Peltier device for temperature control...........46
2-5-3 Charge-coupled-device for imaging process........50
2-5-4 A portable system for AST........................52
Chapter 3 Results and Discussion.......................58
3-1 Capture rate of the Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa , Enterococcus faecalis by using nano-magnetic beads.........................................58
3-2 Bacteria quantification and MIC determination by using resazurin as an indicator..............................60
3-3 Characterization of the microfluidic devices.......62
3-3-1 Results of the measurements of the pumping volume of micropumps and checking the function of two-fold serial dilutions by pneumatic control module...........................62
3-3-2 Temperature profile..............................65
3-3-3 Colorimetric analysis of AST on chip.............67
Chapter 4 Conclusions and future perspectives..........79
4-1 Conclusions........................................79
4-2 Future perspectives................................80
References.............................................82

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